Anti-Mullerian Hormone (AMH), a member of the Transforming Growth Factor (TGF)-beta family, has important roles in normal male and female reproductive development [1]. In addition, AMH has clinical applications in reproductive endocrinology and potentially oncology, which has focused attention on the AMH signal transduction pathway, with the goal of identifying new approaches for therapeutic intervention and diagnostics [2,3]. Like other members of the TGF-beta family, AMH signals by assembling a transmembrane serine/threonine kinase receptor complex of type I and type II components, resulting in the phosphorylation and activation of type I receptor kinase by the constitutively active kinase domain of the type II receptor. The activated type I receptor then phosphorylates the cytoplasmic Smad proteins 1, 5, or 8, which migrate into the nucleus and, in concert with other transcription factors, regulate responsive genes [4,5]. AMHRII, the type II receptor, and AMH, are mutually specific, while ALKs 2, 3 and 6 serve as type I receptors for both AMH and members of the Bone Morphogenetic Protein (BMP) family [6,7]. AMH is translated as a homodimeric precursor, containing an N-terminal pro-region and a smaller C-terminal mature domain. The precursor undergoes an obligatory cleavage at monobasic sites between the two domains, but the pro-region and C-terminal homodimers remain associated in a noncovalent complex. Unlike other TGF-β ligands, the noncovalent complex can bind to AMHRII, which induces dissociation of the pro-region [8]. A similar mechanism has been proposed for the BMP-7 noncovalent complex [9]. A model is presented in FIG. 1, showing processing of AMH, assembly of the AMH receptor signaling complex, and intracellular signaling.
In the male vertebrate embryo, AMH is responsible for the regression of Mullerian ducts, the anlagen of the uterus, Fallopian tubes, and upper part of the vagina. In the adult male, AMH plays a role in Leydig cell differentiation and function [10]. In females, the role of AMH has been predominantly elucidated in rodents, where it has been shown to have an inhibitory effect on primordial follicle recruitment as well as on the responsiveness of growing follicles to Follicle-Stimulating Hormone (FSH) [11,12]. AMH is expressed in Sertoli cells of the fetal and postnatal testis and granulosa cells of the postnatal ovary, whereas AMHRII is expressed in the mesenchymal cells surrounding the Mullerian duct (in both male and females), Sertoli cells, Leydig cells, and granulosa cells. Expression of AMHRII persists in the adult female reproductive tract and has also been detected in the nervous system [13,14,15].
In addition to its role in normal reproductive physiology, AMH is now recognized as an important clinical marker for diagnosing and assessing reproductive disorders in both men and women. In males, serum AMH can be used to assess Sertoli cell function in children with intersex states that can help to distinguish between defects of male sexual differentiation caused by abnormal testicular determination and those resulting from isolated impairment of testosterone secretion or action [16]. In females, the serum AMH level is a reliable marker for the size of the ovarian follicle pool and a predictor of the ovarian response to controlled ovarian hyperstimulation [17]. Furthermore, AMH levels are 2-3 fold higher in women with polycystic ovary syndrome (PCOS) and there is a correlation between the severity of the disease and AMH levels [18]. It has been suggested that the increased follicular growth, which occurs during the early stages of PCOS, may be due to a deficiency of AMH [19], while the follicular arrest observed at later stages could be due to excessive AMH levels [20].
AMH and AMHRII have also been of interest in the field of oncology. AMHRII is expressed on a number of tumors and tumor cell lines [3,21], and AMH has been shown to inhibit the growth of some of these tumors [3]. In addition to developing AMH as a potential therapeutic [3], it has been suggested that agonist antibodies could be generated that bind specifically to AMHRII and trigger the regression of ovarian tumors, by mimicking the ability of AMH to assemble an active receptor signaling complex [22]. Alternatively, antibodies to AMHRII could be coupled to toxins to treat cancers that express AMHRII [23,24].
Various ELISA assays have been developed for detecting AMH and measuring AMH levels in human body fluids [25-27]. Most if not all of these assays employ monoclonal antibodies (mAbs) that detect the pro-region and mature domains. One of the mAbs is used to capture the AMH, while the other is biotinylated and used to detect the captured AMH. While these assays are very sensitive and can detect AMH at low levels in human body fluids, they do not distinguish between uncleaved inactive AMH and the cleaved active noncovalent AMH complex. To date, therefore, all AMH measurements made in normal and disease samples have reported total AMH levels (i.e. uncleaved AMH plus bioactive cleaved AMH) and have provided no information concerning the level of AMH that is active.